Muscle morphogenesis, as it occurs in the avian limb, involves cell migration, splitting of premuscle masses, associations of myogenic cells with connective tissue cells and with matrices deposited by such cells, mutual recognition and adhesion of myogenic cells, myoblast fusion, and elongation of muscle fibers. The molecular mechanisms of these cell-cell and cell-matrix interactions are poorly understood, though it is widely assumed that changes at the cell surfaces of myogenic cells (and their non-myogenic neighbors) as they pass through successive stages of cytodifferentiation will be crucially important. We will use a range of cell culture systems, some of them novel (myoblast suspension cultures; myogenic cells in defined medium) in further developing in vitro assays for morphogenetic processes. Our assay for myoblast attachment and elongation will be used in continuing studies of the mechanism by which the glycoprotein fibronectin mediates these processes. Fibronectin, found in connective tissue matrices and body fluids, has binding sites for collagen, glycosaminoglycans, fibrinogen, other fibronectin molecules, and other ligands, and has been implicated in a variety of biological processes. Limited proteolysis cleaves fibronectin into fragments that retain one or more of the activities listed. The chymotryptic fragments we have prepared from horse serum fibronectin allow the localization of the sites at which fibronectins bind to each other. We will obtain other proteolytic fragments, using other proteases. We will propose and test a model for the arrangement of the different functional sites in the horse serum fibronectin molecule. Oriented fibronectin-containing matrices will be used to test our hypothesis that fibronectin has an organizing function in muscle morphogenesis. In vitro assays for cell aggregation, fusion and attachment to the substratum will be used as part of a monovalent antibody strategy to identify, characterize, and study the function(s) of, other cell surface molecules involved in morphogenesis; activity of antigens in blocking inhibition by monovalent anti-cell surface antibodies will be used in purifying the active molecule(s). Monoclonal antibodies will be obtained against each active molecule so identified.